1. Field of the Invention
The present invention generally relates to enzyme immobilizing carriers and immobilized lipase which are used as a bioreactor, biosensor, and so on to facilitate various biochemical reactions in an industrial field wherein various enzymes are employed as a biological catalyst.
2. Description of the Prior Art
In order to execute various biochemical reactions in an industrial field by using an organic catalyst, for example, an enzyme catalyst, many researches and studies on an immobilized enzyme bioreactor and a biosensor have been actively carried out in recent years. On the same occasion, studies on a carrier for immobilizing enzyme; i.e., an enzyme immobilizing carrier, which is applied to these bioreactor and biosensor, have also been developed. This immobilized enzyme bioreactor includes a column which is filled with an organic catalytic material made of an immobilized enzyme carrier. For the carrier, various materials may be used, for example, high molecular organic materials and inorganic materials such as commonly used porous glass, ceramics, and so on.
Although such inorganic materials have some advantages such as a high mechanical strength, and an excellent thermal and chemical stability during a sterilization treatment, they have the disadvantage that they cannot be repeatly used, due to a poor adsorption force for enzymes. Some conventional arts have been proposed to overcome such adisadvantage. For example, Japanese Patent Publication No.55-32357 and No.56-15231 disclose one typical process where an amount of silane coupling agent introduced with reactive organic radicals are combined with an inorganic carrier such as porous glass, alumina, silica, and so on, and then the enzyme's functional groups are reacted with the reactive organic radicals of the silane coupling agent to realize bonding between the inorganic carrier and the enzyme.
As a typical example of such enzyme immobilized in the inorganic carrier (hereinafter, referred to "immobilized enzyme") including the silane coupling agent, carbohydrate-decomposing enzymes such as .alpha.-amylase, glucoamylase, and so on are known. However, since these enzymes have a low activity; i.e., a poor reactivity with starch or other substrates, compared to the enzyme not immobilized in carrier, a great amount of the immobilized enzyme is required to obtain the same level of reactivity in a non-immobilized enzyme. Accordingly, the immobilized enzyme is not adapted for a producing process in an industrial field. In order to overcome this problem, Japanese Patent Publication No. 5-219952 and 4-51894 disclose a novel carrier and method for immobilizing enzyme into the carrier.
On the other hand, one of typical fat splitting enzyme, lipase, has been rapidly and widely used for various techniques in the field of organic synthesis chemistry such as monoester decomposition, ester synthesis, optical resolution of racemic modification by ester exchange reaction, and so on which is like a splitting function on oil and fat. Nevertheless, it has been only studied and developed as an industrial immobilizing method to physically adsorb a high concentrated lipase solution into diatomaceous earth. In order to produce lipase-adsorbed diatomaceous earth, it is necessary to mix the high concentrated solution of coarse enzymic lipase with diatomaceous earth and then dry the mixture. However, the physical adsorbability of the diatomaceous earth is limited and its adsorbing force is relatively low owing to the physical adsorption. Accordingly, the activity of the lipase-adsorbed diatomaceous earth will gradually decrease by repetition.
Primarily, a main object of using the immobilized enzyme is that although the enzyme is expensive, it can be repeatly used without disposal after only one usage, therefor decreasing production cost and the reacted product is free from any remaining enzymes. In other words, it is desirable to realize a specific immobilized enzyme which can be repeatly used for a long period with the smallest amount possible. As the activity of immobilized enzyme is increased, a scale of reaction apparatus becomes smaller thereby providing the merit of lowering its initial and running cost.